Antibody responses to galectin-8, TARP and TRAP1 in prostate cancer patients treated with a GM-CSF-secreting cellular immunotherapy

A critical factor in clinical development of cancer immunotherapies is the identification of tumor-associated antigens that may be related to immunotherapy potency. In this study, protein microarrays containing >8,000 human proteins were screened with serum from prostate cancer patients (N = 13) before and after treatment with a granulocyte–macrophage colony-stimulating factor (GM-CSF)-secreting whole cell immunotherapy. Thirty-three proteins were identified that displayed significantly elevated (P ≤ 0.05) signals in post-treatment samples, including three proteins that have previously been associated with prostate carcinogenesis, galectin-8, T-cell alternative reading frame protein (TARP) and TNF-receptor-associated protein 1 (TRAP1). Expanded analysis of antibody induction in metastatic, castration-resistant prostate cancer (mCRPC) patients (N = 92) from two phase 1/2 trials of prostate cancer immunotherapy, G-9803 and G-0010, indicated a significant (P = 0.03) association of TARP antibody induction and median survival time (MST). Antibody induction to TARP was also significantly correlated (P = 0.036) with an increase in prostate-specific antigen doubling time (PSADT) in patients with a biochemical (PSA) recurrence following prostatectomy or radiation therapy (N = 19) from in a previous phase 1/2 trial of prostate cancer immunotherapy, G-9802. RNA and protein encoding TARP and TRAP1 was up-regulated in prostate cancer tissue compared to matched normal controls. These preliminary findings suggest that antibody induction to TARP may represent a possible biomarker for treatment response to GM-CSF secreting cellular immunotherapy in prostate cancer patients and demonstrates the utility of using protein microarrays for the high-throughput screening of patient-derived antibody responses.     

The Empirical Analysis of Cigarette Tax Avoidance and Illicit Trade in Vietnam, 1998-2010

Illicit trade carries the potential to magnify existing tobacco-related health care costs through increased availability of untaxed and inexpensive cigarettes. What is known with respect to the magnitude of illicit trade for Vietnam is produced primarily by the industry, and methodologies are typically opaque. Independent assessment of the illicit cigarette trade in Vietnam is vital to tobacco control policy. This paper measures the magnitude of illicit cigarette trade for Vietnam between 1998 and 2010 using two methods, discrepancies between legitimate domestic cigarette sales and domestic tobacco consumption estimated from surveys, and trade discrepancies as recorded by Vietnam and trade partners. The results indicate that Vietnam likely experienced net smuggling in during the period studied. With the inclusion of adjustments for survey respondent under-reporting, inward illicit trade likely occurred in three of the four years for which surveys were available. Discrepancies in trade records indicate that the value of smuggled cigarettes into Vietnam ranges from $100 million to $300 million between 2000 and 2010 and that these cigarettes primarily originate in Singapore, Hong Kong, Macao, Malaysia, and Australia. Notable differences in trends over time exist between the two methods, but by comparison, the industry estimates consistently place the magnitude of illicit trade at the upper bounds of what this study shows. The unavailability of annual, survey-based estimates of consumption may obscure the true, annual trend over time. Second, as surveys changed over time, estimates relying on them may be inconsistent with one another. Finally, these two methods measure different components of illicit trade, specifically consumption of illicit cigarettes regardless of origin and smuggling of cigarettes into a particular market. However, absent a gold standard, comparisons of different approaches to illicit trade measurement serve efforts to refine and improve measurement approaches and estimates.     

Diet of the Indochinese silvered langur (Trachypithecus germaini) in Kien Luong Karst area,Kien Giang Province

The Indochinese silvered langur (Trachypithecus germaini) is distributed to the west of Mekong River in Cambodia, Lao PDR, Thailand and Vietnam. During a two‐year study, from May 2014 to May 2016, we collected 320.44 hr of behavior, with 17,040 feeding bouts recorded (142 hr) for T. germaini on Chua Hang Karst Mountain, Kien Luong District, Kien Giang Province, Vietnam. Feeding accounted for 45% of the Indochinese silvered langurs’ activity budget. The plant diet of the Indochinese silvered langurs was principally composed of young leaves (58%), followed by mature leaves (9.5%), fruits (22.7%), flowers (4.7%), buds (3.3%), petioles (1.2%), and other (0.5%). A total of 58 plant species were fed on by the silvered langurs, and leaves of eight species (Phyllathus reticulatus, Ficus rumphii, Ficus tinctoria, Ficus microcarpa, Cayratia trifolia, Streblus ilicifolia, Combretum latifolium, and Streblus asper) were fed on throughout the year. P. reticulatus was most frequently eaten (13.9% feeding time, n = 1,733). Food selection differed significantly between months and seasons. The Indochinese silvered langurs ate 27 plant species in the wet season compared with 23 plant species in the dry season. Leaf chemical composition of two food categories, 16 eaten species (with 10 most frequently consumed species and six least consumed species), and four noneaten species, were analyzed. Feeding samples from eaten species in the Indochinese silvered langurs's diet contained lower amounts of condensed tannin, lignin, protein, ash, and lipids, but a higher amount of total sugar compared with samples from noneaten species. Furthermore, the most frequently consumed species contained lower amounts of lignin compared with the less frequently consumed species. Using a generalized linear model with five variables, including neutral detergent fiber (NDF), total sugar, lignin, lipid, and calcium (Ca) indicated that NDF positively correlated and lignin content negatively correlated with feeding records in the diet of these langur.     

Antilipolytic activity of viprostol, a transdermally active antihypertensive PGE2 analog

Viprostol, a novel prostaglandin E2 congener, was assessed for in vitro antilipolytic activity in the spontaneously obese rat. In isolated epididymal adipocytes, viprostol exhibited a dose-dependent inhibition of catecholamine-stimulated lipolysis at concentrations ranging from 10 microM to 1 mM, but was ineffective at lower concentrations. Additionally, viprostol exhibited approximately 50% of the antilipolytic activity of naturally-occurring PGE1 and PGE2 at similar concentrations, but was as potent as PGF2 alpha. At 10 microM, viprostol inhibited maximum catecholamine-stimulated lipolysis by approximately 35% of the total, hormone-stimulated glycerol release. The results of these experiments indicate that viprostol exhibits antilipolytic activity in vitro, but is less potent than the naturally-occurring PGE's to which it is most closely related structurally.     

Drought Stress and Elevated CO(2) Effects on Soybean Ribulose Bisphosphate Carboxylase Activity and Canopy Photosynthetic Rates

Soybean (Glycine max [L.] cv Bragg) was grown at 330 or 660 microliters CO₂ per liter in outdoor, controlled-environment chambers. When the plants were 50 days old, drought stress was imposed by gradually reducing irrigation each evening so that plants wilted earlier each succeeding day. On the ninth day, as the pots ran out of water CO₂ exchange rate (CER) decreased rapidly to near zero for the remainder of the day. Both CO₂-enrichment and drought stress reduced the total (HCO₃⁻/Mg²⁺-activated) extractable ribulose-1,5-bisphosphate carboxylase (RuBPCase) activity, as expressed on a chlorophyll basis. In addition, drought stress when canopy CER values and leaf water potentials were lowest, reduced the initial (nonactivated) RuBPCase activity by 50% compared to the corresponding unstressed treatments. This suggests that moderate to severe drought stress reduces the in vivo activation state of RuBPCase, as well as lowers the total activity. It is hypothesized that stromal acidification under drought stress causes the lowered initial RuBPCase activities. The K_m(CO₂) values of activated RuBPCase from stressed and unstressed plants were similar; 15.0 and 12.6 micromolar, respectively. RuBP levels were 10 to 30% lower in drought stressed as compared to unstressed treatments. However, RuBP levels increased from near zero at night to around 150 to 200 nanomoles per milligram chlorophyll during the day, even as water potentials and canopy CERs decreased. This suggests that the rapid decline in canopy CER cannot be attributed to drought stress induced limitations in the RuBP regeneration capability. Thus, in soybean leaves, a nonstomatal limitation of leaf photosynthesis under drought stress conditions appears due, in part, to a reduction of the in vivo activity of RuBPCase. Because initial RuBPCase activities were not reduced as much as canopy CER values, this enzymic effect does not explain entirely the response of soybean photosynthesis to drought stress.     

l-Phenylalanyl-l-Glutamate-Stimulated, Chloride-Dependent Glutamate Binding Represents Glutamate Sequestration Mediated by an Exchange System

Stimulation of glutamate binding by the dipeptide L-phenylalanyl-L-glutamate (Phe-Glu) was inhibited by the peptidase inhibitor bestatin, suggesting that the stimulation was caused by glutamate liberated from the dipeptide and not by the dipeptide itself. It further suggests that this form of glutamate binding should be reinterpreted as glutamate sequestration and that stimulation of binding both by dipeptides and after preincubation with high concentrations of glutamate is likely to be due to counterflow accumulation. Several other criteria indicate that most of glutamate binding stimulated by chloride represents glutamate sequestration: Binding is reduced when the osmolarity of the incubation medium is increased, when membranes incubated with [3H]glutamate are lysed before filtration, and when membranes are made permeable by transient exposure to saponin. Moreover, dissociation of bound glutamate after a 100-fold dilution of the incubation medium is accelerated about 50 times by the addition of glutamate to the dilution medium. This result would be anomalous if glutamate were bound to a receptor site; it suggests instead that glutamate is transported in and out of membrane vesicles by a transport system that preferentially mediates exchange between internal and external glutamate. Glutamate binding contains a component of glutamate sequestration even when measured in the absence of chloride. Sequestration is adequately abolished only after treating membranes with detergents; even extensive lysis, sonication, and freezing/thawing may be insufficient.     

小麦叶绿体中CTK结合蛋白的结合活性研究

叶绿体中存在着与细胞分裂素(CTK)专一结合的蛋白质。这一蛋白与6-苄氨基嘌呤(6 BA)的亲和力很强,解离常数达3.7×10~(-8)mol/L。最大结合量为10.7 pmol 6 BA/mg蛋白,Seatchard分析表明只有一类结合位点。不同的叶绿体纯化步骤对CTK结合蛋白的活性有不同的影响,分离步骤少而快速的差速离心法可以得到具有较高结合活性的叶绿体。叶绿体经分离纯化后,低温保存时的结合活性较稳定,-20℃以下可以较长期保存。用EDTA预处理叶绿体,不降低CTK结合蛋白对6 BA的结合活性,而用高浓度的NaCl处理,可以使叶绿体结合6BA的能力明显下降。这说明EDTA不能使CTK结合蛋白从叶绿体膜系统表面解离,而高浓度NaCl则有这种可能性。     

Immunochemical characterization of two antigenic sites on human apolipoprotein A-I; localization and lipid modulation of these epitopes

Two monoclonal antibodies, A17 and A30, were raised against human apolipoprotein A-I (apo A-I). They were studied by competitive inhibition of 125I-labeled HDL3 with HDL subfractions, delipidated apo A-I, and complexes of dimyristoylphosphatidylcholine (DMPC) containing apo A-I and apo A-II. Immunoblotting located the A17 antibody on CNBr fragment 4 of apo A-I and the A30 antibody on CNBr fragment 1. The A17 antigenic determinant was expressed identically in all HDL subclasses, on delipidated apo A-I as well as all on the DMPC-apo A-I and DMPC-apo A-I/apo A-II complexes. In contrast, the apparent affinity constant of the A30 antibody for delipidated apo A-I was about 30-times less than for HDL3 or for apo A-I/apo A-II-phospholipid complexes. These data suggest that the association of apo A-I with phospholipids improves the reactivity of the A30 monoclonal antibody towards apo A-I, and that this antigenic determinant has a different conformation in delipidated apo A-I compared to apo A-I complexed with phospholipids. Turbidimetric and fluorescence experiments monitoring the phospholipid-apo A-I association in the presence and in the absence of the A17 and A30 antibodies were consistent with the competition experiments carried out by solid phase radioimmunoassay (RIA). After reaction of apo A-I with the A30 antibody, we observed an enhancement of the degradation kinetics of large multilamellar vesicles (LMV), while the A17 antibody did not have a significant effect. Calcein leakage experiments carried out below the transition temperature of DPPC showed an enhancement of the degradation kinetics with both monoclonal antibodies, while the phase-transition release was independent of the reaction of apo A-I with the monoclonal antibodies. These data therefore suggest the existence of at least two different types of epitope on apo A-I, which might account for the differences in immunological reactivity of apo A-I that is either delipidated or present on HDL.